Hematopoietic stem/progenitor cells represent a very small fraction of cells in the bone marrow which have the capacity for both self- renewal and differentiation along multiple paths ultimately progressing to form all of the cellular elements of peripheral blood. Much remains to be understood about these cells and the patterns of gene expression that characterize them. The current proposal describes an approach to the isolation of highly enriched murine hematopoietic stem/progenitor cells and synthesis of cDNA which accurate represents the genes expressed by these cells in sufficient quantities to permit further study. Amplified cDNA from various populations of stem/progenitor cells will be used as substrates for representational difference analysis (RDA), a PCR- based subtractive hybridization technique for identifying genes that are expressed differentially between populations. Fragments of genes isolated in this way, that are confirmed to be preferentially expressed by primitive stem/progenitor cells, will be compared to sequence databases. Novel gene fragments will be used to isolate full length cDNAs from stem/progenitor cell cDNA libraries. These cDNAs will be assessed for the presence of characteristic structural motifs which will direct further experiments. The patterns of expression of genes identified will be examined. The central hypothesis is that hematopoietic stem/progenitor cells are characterized by unique patterns of gene expression which define their phenotype. The goal is to apply molecular techniques to the identification of genes of particular relevance to stem cell growth and differentiation for the purpose of advancing cell purification, ex vivo expansion and genetic manipulation.